Humans are exposed to polycyclic aromatic hydrocarbons (PAHs) from various occupational, environmental, medicinal and dietary sources. The measurement of specific PAH metabolites, particularly 1-hydroxypyrene, in human urine treated with deconjugating enzymes (e.g. β-glucuronidase) has been extensively used as a means of assessing recent exposure to PAHs. We have examined pyrene metabolites in human urine prior to enzymatic deconjugation in order to determine the relative proportions of conjugated and unconjugated pyrene metabolites. The analytical method utilized immuno-affinity chromatography, high performance liquid chromatography (HPLC) and the complementary techniques of synchronous fluorescence spectroscopy (SFS) and gas chromatography-mass spectrometry (GC-MS) to measure pyrene-containing metabolites. SFS analysis of immunoaffinity-purified urine samples showed fluorescence spectra characteristic of the pyrene moiety (using wavelength differences of 34 nm, 54 nm and 102 nm). These spectra are produced by several PAHs containing the pyrene moiety. HPLC analysis with fluorescence detection indicated that the major fluorescent metabolite in immunoaffinity-purified urine was much more polar than simple hydroxylated metabolites of pyrene (1-hydroxypyrene) or benzo[a]pyrene (benzofa] pyrene-diols or -tetrols). Following digestion with β-glucuronidase, this metabolite co-chromatographed with authentic 1-hydroxypyrene and exhibited fluorescence spectra characteristic of 1-hydroxypyrene, suggesting that the major metabolite was a glucuronide conjugate of 1-hydroxypyrene. This was subsequently confirmed by GC-MS analysis of trimethylsilyl derivatives of the major metabolite; both 1-hydroxypyrene and glucuronic acid were detected independently as derivatized products. Since 1-hydroxypyrene glucuronide is approximately 5-fold more fluorescent than 1-hydroxypyrene, it may provide a more sensitive biomarker for assessing exposure to pyrene in mixtures of PAHs.
ASJC Scopus subject areas
- Cancer Research